Similar embryotoxic effects of sera from infertile patients and exogenous interferon-gamma on long-term in-vitro development of mouse embryos

Abstract

Circulating embryotoxic factors could be responsible for reproductive failures observed in patients suffering from recurrent spontaneous abortions (RSA) and endometriosis. The mouse bioassay has been widely used to detect such factors, since sera from these patients inhibit early embryonic development. This bioassay consists in the in-vitro culture of two-cell mouse embryos in the presence of different sera up to the blastocyst stage (72 h of culture). In the present study experiments were performed over long culture times (3-7 days), from two-cell to spreading stages, to determine the in-vitro effect of sera obtained from RSA or endometriosis patients, as well as the effect of interferon (INF)-gamma on embryo development. An embryotoxicity cut-off value of 45% blastocyst formation was established using control sera. When development to the blastocyst stage was considered only 25% of RSA and 20% of endometriosis sera were embryotoxic. However, all RSA sera significantly inhibited hatching (P < 0.05) and spreading stages (P < 0.01). IFN-gamma (10 micrograms/ml) (P < 0.001) did not impair early embryo development, but significantly inhibited blastocyst spreading. These observations suggest that culture to advanced embryonic stages increases the sensitivity of the bioassay and that IFN-gamma alters in-vitro peri-implantation mouse embryo development.

Exogenous interferon-γ alters murine inner cell mass and trophoblast development. Effect on the expression of ErbB1, ErbB4 and heparan sulfate proteoglycan (perlecan)

“…To examine the effect of IFN-g, a group of embryos (experimental group) was cultured at the lowest concentration of cytokine producing a significant inhibition of outgrowth (Cameo et al 1999). Human recombinant IFN-g (3 £ 10 5 iu/ml; Boehringer Ingelheim, Vienna, Austria) was added to the medium at different culture times: day 0, 3 or 4 of culture.…”

“…IFN-g receptors have been found in mouse oocytes and pre-implantation embryos (Truchet et al 2001) and IFN-g level is increased in sera of women suffering from spontaneous miscarriage (Jenkins et al 2000). In a previous study we demonstrated that human recombinant interferon-g (hIFN-g) added to the culture medium inhibited mouse blastocyst development impairing the outgrowing stage, mimicking the effect of embryo exposure to sera from women suffering from recurrent spontaneous abortions (Cameo et al 1999). Given that normal embryo outgrowth is a prerequisite for successful implantation, these results suggest that embryos exposed to high concentrations of the cytokine could have an altered spatio-temporal expression of molecules that are crucial for the implantation process, such as growth factors and their receptors.…”

Luteal phase serum cell-free DNA as a marker of failed pregnancy after assisted reproductive technology

“…About 24% (8,10) to 70% (2) of the sera of RSA patients and 0% (9) to 100% (7) of the endometriosis patients were found to be embryotoxic. Recently, this laboratory (11) and others (12) demonstrated the presence of DNA-damaging embryotoxic factors in patients that failed to achieve pregnancy and embryo implantation after assisted reproductive technologies (ART).…”

“…Embryotoxic factors have been detected in the sera of patients with recurrent spontaneous abortions (RSA) or endometriosis (1)(2)(3)(4)(5)(6)(7)(8)(9)(10). About 24% (8,10) to 70% (2) of the sera of RSA patients and 0% (9) to 100% (7) of the endometriosis patients were found to be embryotoxic.…”

Sperm Apoptosis in Nonpregnant Lutealphase Sera After in Vitro Fertilization as Assessed by Comparative Genomic Hybridization

“…The serum factors were not involved in the pregnancy losses of biochemical pregnancies as shown by the rather similar fluorescent intensities of the biochemical versus clinical pregnancy results. Other researchers have suggested that the embryotoxic factors present in the sera of RSA and endometriosis patients are either a Th1-type cytokine, IFN-g [2,7], cytokeratin inhibitors that damage cellular cytoskeleton [4], or Fas-mediated immunoglobulins that fragment DNA [1,10]. The embryotoxic factors involved in pregnancy losses after assisted reproductive technologies procedures remain unknown.…”